Plants require nitrogen during their vegetative and reproductive growth phases. Nitrogen is made available to the plant through soil mineralization, the application of nitrogen fertilizer, or both. It has been estimated, however, that between 50 and 70 percent of nitrogen applied to crops is lost from the plant-soil system [Peoples, M. B. et al., “Minimizing Gaseous Losses of Nitrogen,” In Nitrogen Fertilizer in the Environment (Bacon, P. E., ed.) Marcel Dekker, pp. 565-606 (1995)]. Nitrogen is one of the most expensive plant nutrients to supply, nitrogen fertilizer is not always available at a reasonable cost, and excessive application of nitrogen fertilizer can result in environmental challenges. Corn is an example of an agronomically important plant that often requires nitrogen fertilizers to perform at its genetic potential.
In plant cells, NAGK (N-Acetyl Glutamate Kinase) carries out the second step in a biosynthetic pathway that leads to the production of arginine. Importantly, several plant and bacterial NAGK enzymes are known to be inhibited by high concentrations of arginine (Bourrellier, 2009; Llacer, 2008; Lohmeier-Vogel, 2005; Fernandez-Murga, 2004), which suggests that NAGK plays a role in arginine regulation in plants. Additionally, it has been demonstrated that the plant P-II protein can mitigate the arginine-based inhibition of NAGK. P-II is known to play a key role in regulation of carbon and nitrogen flow in plants (Bourrellier, 2009; Llacer, 2008).
One strategy to upregulate the arginine biosynthetic pathway in plants is to reduce or eliminate the inhibition of NAGK by arginine. This can be achieved by heterologous expression of an arginine-insensitive NAGK enzyme in plants. Here, we describe the overexpression and enzymatic characterization of an arginine-insensitive NAGK isolated from a bacterial strain.